Currently in drug discovery and development and in general biological research, methods and apparatus for accurately performing cell-based assays are used. Cell-based assays are advantageously employed for assessing the biological activity of chemical compounds.
In assessing the biological activity of chemical compounds, there is a need to quickly and inexpensively screen large numbers of chemical compounds. This need has arisen in the pharmaceutical industry where it is necessary to test chemical compounds for activity against a variety of biochemical targets, for example, receptors, enzymes and nucleic acids. These chemical compounds are collected in large libraries, sometimes exceeding one million distinct compounds. The use of the term chemical compound is intended herein to be interpreted broadly so as to include, but not to be limited to, simple organic and inorganic molecules, proteins, peptides, nucleic acids and oligonucleotides, carbohydrates, lipids, or any chemical structure of biological interest.
In the field of chemical compound screening, cell-based assays are run on populations of cells.
International patent application WO 99/47963 describes a translocation assay in which two or more cell species, for example a biological cell nucleus and a biological cell transcription factor protein, are fluorescently labelled. In the assay, images are acquired including both the nucleus and the transcription factor species. The images are processed such that a co-localisation of the two species may be determined in order to analyse a migration of the transcription factor within the cell. For the course of time of the translocation assay, the cell nucleus is fluorescently labelled with a nuclear dye.
International patent application WO 03/031612 describes a process for determining a phase of a biological cell cycle by analysing nucleic acid, for example DNA, of cell nuclei of the cells as a function of time. Images of the cells are recorded over the course of the assay and, in order to analyse the cell nuclei using these images, the nuclei are stained using a nuclear dye
The staining of cellular nucleic acids, for example DNA, with a nuclear dye such as Hoechst (propidium iodide) or DRAQ5, is used to identify a spatial definition of a cell nucleus, from which to form a nuclear mask. The properties of the cell may be determined by measuring characteristics of the cell inside and outside the nuclear mask area. However, the nuclear dye is destructive to the structure of the nucleic acid and hence multiple samples of cells are required to study the progression of biological activity over time. In the case where many hundreds of samples are to be analysed, this deterioration of the nucleic acid can become rate limiting.
The constituent molecules of such nuclear dyes bind to the molecule of the nucleic acid molecule by intercalating between the base pairs of the nucleic acid. This intercalation, however, over the course of an experiment having an extended time period proves to be toxic to the nucleic acid. The toxicity results in damage to the structure of the nucleic acid leading to mutations in the genetic code. These mutations interfere with the correct functioning of the cell and in the case of screening chemical compounds for their effects on biological cell systems, introduces undesired errors into results data being collected.
It is an object of the present invention to address the problems in the prior art relating to the use of a marker such as a nuclear dye.